In renal failure, loss of kidney function results in uremic syndrome, characterized by the accumulation of salts, water and toxic breakdown products of protein metabolism in the blood. There are approximately one million people worldwide with chronic kidney failure, with an annual growth rate of around 9%. Current therapies are based on conduction (diffusion—passive transport of solutes from blood to dialysate across a dialysis membrane), convection (ultrafiltration—simultaneous transport of solvent and solute from the blood compartment to the dialysate compartment across a dialysis membrane) and adsorption (protein adsorption depends on how hydrophobic the membranes are) strategies. Dialysis is a therapy which eliminates the toxic wastes from the body due to kidney failure. There are two main types of dialysis: hemodialysis and peritoneal dialysis. The majority (85%) of end stage renal disease (ESRD) patients are treated using a technique called hemodialysis which uses a passive diffusion strategy.
Hemodialysis entails the re-routing of blood from the body to a filter made of plastic capillaries, for example, passing blood through an artificial kidney, where uremic toxins, such as salts and urea (low molecular weight molecules), diffuse across a semipermeable membrane into isotonic dialysate, resulting in reduced toxin concentration in the blood. The blood is purified when the waste products diffuse from the blood across the membrane of these tiny capillaries. The blood is then returned to the body, usually via a vein in the arm. However, as toxic molecules increase in size, their ability to be removed by diffusion decreases. Typically, only 10% to 40% of larger molecules, called middle molecular weight molecules, are removed during a dialysis session. Consequently, these toxins reach abnormally high levels and begin to damage the body over time. The inefficient removal of uremic toxins, such as middle molecule toxins (e.g. β-2-microglobulin) and phosphate represent significant limitations of current renal dialysis technology. Currently, to achieve minimum adequate removal of urernic toxins, manufactures and nephrologists are attempting to increase the surface area of the artificial kidneys and prolong the patient treatment times. However, as blood priming is required for dialysis, there is a limit to the surface area of an artificial kidney. The limit will be reached when the blood priming volume (which goes to waste) exceeds the human blood regeneration time. Also, increasing therapy times reduces the quality of life of a patient and increases the medical and ancillary staff requirements. Often, patients suffer from side effects, or morbidity, due to the inability of the hemodialysis procedure to completely replace the function of a normal living kidney.
In peritoneal dialysis, the body's own membrane is used as a filter, and the fluid drained in and out of the abdomen replaces the kidneys in getting rid of toxins. However, this technique may result in peritonitis and membrane failure.
In healthy individuals, the kidney functions to remove excess water, salts and small proteins from the blood circulation. Nitrogenous wastes removed by the kidney include urea, the final metabolic destiny of excess dietary nitrogen, creatinine which is produced during muscle activity, and uric acid (an endpoint product of nucleotide metabolism). Current renal dialysis technology relies on equilibrium/diffusion principles and transmembrane pressure to remove nitrogenous wastes, salts and excess water from the bloodstream of patients experiencing chronic or acute renal failure. Current dialysis technologies suffer from sub-optimal biocompatibility of the dialysis membranes used, the inadequacy of existing technology in the removal of some solutes such as phosphates, and poor removal of low molecular weight proteins such as beta-2 microglobulin.
Beta-2-microglobulin (β2m) associated ainyloidosis affects long term dialysis patients. Under normal physiological conditions, free circulating β2m can be found in plasma at low concentrations (1–3 mg/L). However, this level can be up to fifty times higher in long term dialysis patients. In healthy individuals, glomerular filtration and catabolism in the proximal renal tubule effectively remove β2m. The impairment of renal function often leads to the retention of β2m and subsequent increase in its circulating concentration. Further, current dialysis treatments are unable to efficiently remove β2m at a sufficient rate leading to its accumulation and deposition as part of amyloid fibrils in various niusculoskeletal structures. These amyloid deposits are predominantly osteoarticular and are associated with various clinical manifestations such as carpal tunnel syndrome, joint pain and stiffliess, bone cysts, pathological fracture and soft tissue masses. The clinical problems associated with β32m amyloidosis constitute a major cause of morbidity in long term dialysis patients.
Beta-2 microglobulin is an 11.9 kDa non-glycosylated protein comprising a polypeptide chain of 99 amino acid residues. It is encoded by a single gene on chromosome 15, and is synthesized with an 18 residue signal peptide. 132m is ubiquitously expressed on the surface of all nucleated cells where it functions as the light chain of HLA class I molecule via a non-covalent association with the heavy chain and is required for transport and expression of the complex at the cell surface. It has also been shown to have amino acid sequence homology with the constant domain of IgG(CH3) and the alpha-3 domain of the heavy chain HLA class I. Several isoforms of β2m have been described, the native β2m and more acidic variants which are found in long term dialysis patients, possessing isoelectric points of 5.7 and 4.8 to 5.3 respectively.
Existing renal dialysis modalities employ diffusive and/or convective means to remove contaminating molecules from blood. Blood is passed through a disposable cartridge where the blood travels in one direction and dialysate, separated from the blood stream by a semi-permeable membrane, flows in the opposing direction.
Diffusive removal means that when blood is dialysed against a physiological dialysate solution, contaminant molecules present in blood, but absent from the dialysate, diffuse down their concentration gradient, out of the blood stream and into the dialysate stream. This is the mode of contaminant removal employed in ‘conventional renal dialysis’. Dialysis of this type is usually adequate for the removal of low molecular weight solutes, but is entirely inadequate for the removal of larger blood components or contaminants like β2 microglobulin.
Convective removal means that blood is processed over a membrane at a pressure sufficient to force fluid (but not cells) through the membrane. This is referred to as hemofiltration, and allows removal of blood contaminants that are carried out by the bulk flow of fluid from the blood. Some fluid removal is the balanced by the infusion of a substitution solution into the patient to maintain correct body fluid balance.
Diffusive and convective means of solute removal may be combined in the dialysis mode known as hemodiafiltration, in which blood, maintained at a relatively high pressure, is dialysed against a dialysate solution maintained at a lower pressure. Solutes are able to diffuse across the semi-permeable membrane, while the simultaneous bulk fluid removal from blood, induced by transmembrane pressure, adds to the rate of solute removal. In HDF, a substitution solution is also used to maintain fluid balance, and may be added to blood before or after processing in the dialysis cartridge.
Gradiflow™ is a membrane-based preparative electrophoresis technology developed by Gradipore Limited (Australia) in which separations are achieved using the dual strategy of molecular charge and size. The distinguishing features of this technology are a set of hydrogel membranes and the application of an electrical potential across these membranes to drive a separation. The use of these features allows the selective removal of contaminants or a product from complex starting materials, which has been demonstrated in a number of protein purifications.